1. Field of the Invention
This invention relates to immunodiagnostic assay techniques and to methods for producing solid carriers which are useful in immunoassays. More particularly, it relates to a method for applying materials to solid carriers such as polystyrenes, polyethylenes and other plastics, woven or non-woven materials, cellulosics such as paper and wood or glass substances, so that the subsequent use of the solid carrier in an immunoassay results in higher sensitivities than are obtained without such material. Still more specifically, it relates to the introduction of a polymeric material onto a solid carrier to result in a carrier which gives very sensitive results upon the subsequent use of that material in an immunoassay measurement.
2. Description of Relevant Literature and Prior Art
Many differing prior art techniques have been used to conduct immunoassays. These techniques have utilized a wide variety of particular indicators, generally called markers, tags, tracers or labels, in the subsequent measurement of an analyte. In most immunoassays, there is a general requirement for the ultimate separation of a component which is labeled or tagged with an indicator from that same component in unlabeled form. A solid support carrier having the appropriate immunological materials attached is used to facilitate that separation.
The well-known sandwich assay for detecting or determining either an antigen or antibody in a sample is illustrative of immunoassays with which the present invention is useful. Assuming, for purposes of illustration only, that it is the antigen which is the analyte to be determined, a capture antibody is either adsorbed or covalently bonded to a solid carrier either directly or through an intermediate bridging layer. With regard to the latter, see U.S. Pat. No. 4,001,583 issued Jan. 4, 1977 to M. James Barrett relating to the use of glutaraldehyde as the means by which the bridging layer is covalently bound to subsequent proteinaceous materials. A biological liquid or a standard solution containing the antigen (or not containing it as the case may be) is incubated in the presence of the carrier/antibody composite for a suitable period of time during which the free antigen reacts with the capture antibody to form a complex on the composite. The complex is then contacted with a labeled component (usually the corresponding antibody) capable of binding to the sought antigen. This labeled component is generally described in the art as the "indicator antibody" because it contains the label which is subsequently used to measure the amount of labeled antibody bound to the antigen (or analyte) sought in the sample.
As indicated previously, many different indicators have been employed in the art. Tracers such as enzymes for enzyme immunoassay (EIA or ELISA) and radioactive labels for the so-called RIAs (radio immunoassays), and fluorescent and bioluminescent materials have all been employed. The art has discovered that for most analytes the best sensitivities are obtained generally from radio-immunoassays or fluorescent assays. Enzyme immunoassays give somewhat lower sensitivities limiting their use. From a practical point of view, those analytes that are present in very small quantities, say of the order of 10 nanograms and below, are most effectively measured utilizing radioimmunoassay as the optimal technique with fluorescent assays being suitable in some analytes. The art has not been favorably disposed to using enzyme immundassays to detect these materials because of the difficulty of detecting concentrations below 10 nanograms. It would be very desirable indeed to provide a technique for obtaining sensitivities from an EIA or ELISA format which approach those of RIA, and indeed for even improving the sensitivities of RIA and other highly sensitive techniques.
One of the characteristics of the immunoassay techniques which tends to prevent obtaining sensitivities that are theoretically possible, relates to the ability of the tag to be detected at low levels in the presence of background activity caused by materials which are non-specifically bound to the capture molecule (antibody, antigen, etc.) or to the carrier itself. Since the RIA and fluorescent assays are inherently more sensitive than EIA, this drawback is less of a problem for them. But it can be seen that successful efforts to improve sensitivity would convert low sensitivity methods such as EIA or ELISA into those having sensitivities approaching or surpassing RIA. The art, as stated previously has attempted to improve this characteristic for all techniques by interposing an intermediate layer of a substance, such as glutaraldehyde, for example, onto the solid carrier prior to the application of the capture molecule. The effect of the glutaraldehyde placement is to bind the capture molecule covalently, i.e. more tightly and presumably in large quantities to the carrier. The latter characteristic would tend to obscure more of the carrier and make it unavailable for non-specific binding subsequently by the indicator molecule.
Similarly, other coating substances have been applied to the solid support in an effort to enhance attachment of the capture molecule. Bovine Serum Albumin has been used to form a physical absorbing layer while various polypeptides such as polyphenylalanine-lysine subsequently activated with glutaraldehyde, as shown in U.S. Pat. No. 4,657,873, have been used as plastic coating materials for covalent bonding to capture molecules.